Spinal cord injury (SCI) is a devastating condition that affects active individuals of all ages for whom treatment options remain inadequate. Two well-established injury mechanisms are reactive oxygen species (ROS)-mediated oxidative damage to lipids and proteins;and the activation of calcium-dependent proteases (calpains) and consequent breakdown of a number of proteins, including cytoskeletal proteins. Despite the progress made in understanding the cellular and molecular mechanisms of neurodegeneration following acute SCI, treatment of this condition has remained a challenge. Therapeutic intervention against ROS mediated oxidative damage using the antioxidant compound Tempol at short time points following acute SCI has been shown to be at least partially neuroprotective. To achieve a greater protective effect following SCI, it would be advantageous to combine more than one therapeutic approach. The overall aim is to systematically block post-traumatic neurodegeneration following SCI by scavenging ROS and inhibiting calpain-mediated proteolytic degradation of the neuronal cytoskeleton. Aim 1 will test the hypothesis that inhibition of calpain, using a novel drug with high cell permeability and blood-brain barrier penetrability, will decrease cytoskeletal damage to the neurons and promote neuroprotection in acute SCI. The rat PSI T10 spinal cord contusion injury model will be used in Aims 1 and 2. Shams, injured rats with vehicle, and injured rats treated with critically selected doses of the calpain inhibitor will be compared at specific time points following injury. Calpain-mediated cytoskeletal breakdown will be assessed using an immunochemical spectrin breakdown assay as an indirect measure of calpain activity. Tissue sparing, functional and locomotor recovery will also be compared among the treatment and control groups. Aim 2 will test the hypothesis that a combination of antioxidant therapy with calpain inhibition will produce an additive and/or synergistic protective effect after SCI. The optimal dosing paradigm for the antioxidant Tempol as previously determined and for the novel calpain inhibitor as determined in Aim 1 will be used in this combinatorial approach. In addition to the methodology used in Aim 1, oxidative damage will be assessed using immunochemical approaches including slot blots, Western blots and immunohistochemistry. The objective of this project is to develop a combinatorial therapeutic strategy for the treatment of SCI in a rat model. The findings of this study will provide a foundation for the design of future translational studies in human SCI.